The present invention relates to compounds useful as enzyme inhibitors, in particular as inhibitors of enzymes involved in the transfer of phosphoryl groups and, especially as inhibitors of polymerases. The invention further relates to pharmaceutical compositions containing such compounds, and to their use in the treatment of viral infections.
Polymerases are the enzymes which catalyse the formation of phosphodiester bonds in RNA and DNA. They play an essential role in viral replication and, therefore, are an important target in the fight against viral diseases such as human immunodeficiency virus (HIV), hepatitis, and poliomyelitis.
U.S. Pat. No. 5,475,109 describes dioxobutanoic acids substituted with piperidine or similar N-substituted saturated cycloalkyls as inhibitors of the cap-dependent endonuclease of influenza virus.
The present inventors have discovered that a range of diketoacids have utility as enzyme inhibitors and, in particular, as polymerase inhibitors and more particularly as inhibitors of hepatitis C NS5 RNA-dependent RNA polymerase, HBV DNA-dependent RNA polymerase and HIV DNA-dependent DNA polymerase. Their investigations indicate that these compounds may act by interfering with the binding of phosporyl groups at the active site of the enzyme and may, therefore, have broad application in inhibiting enzymes involved in the transfer of phosphoryl groups.
According to a first aspect of the present invention there is provided a compound of formula A shown below. This compound is suitable for therapeutic use, for instance as an enzyme inhibitor. 
Optionally, the compound may be in the form of a pharmaceutically acceptable salt or ester, which can be hydrolysed in vivo to the corresponding diketoacid.
In formula A, the group R is an organic moiety which contains from 2 to 24, preferably 4 to 20, most preferably 6 to 17 carbon atoms in total. R includes an optionally substituted cyclic or heterocyclic group in which the atom directly bonded to the adjacent carbonyl in the diketoacid is part of the ring structure. Preferably, this atom is a carbon atom.
The ring which is thus bonded to the carbonyl group is preferably a 3 to 8 membered ring, particularly a 4 to 6 membered ring.
Thus, for example, R may be selected from:
(i) optionally substituted aromatic groups, especially those including six membered rings, such as phenyl and naphthyl;
(ii) optionally substituted heteroaryl groups especially those including five and six membered rings such as thiophene, pyrrole, furan, imidazole, pyridyl, pyrimidyl, and pyridazyl; the heteroaryl ring may, optionally be fused to another ring;
(iii) optionally substituted cycloalkyl groups, especially those including five or six membered rings such as cyclopentyl, cyclohexyl and adamantyl;
(iv) optionally substituted cycloalkenyl groups, especially those including five or six numbered rings such as cyclohexenyl, cyclopentenyl;
(v) optionally substituted cyclic heteroalkyl groups, especially those including five or six numbered rings such as piperidyl, pyrrolidyl, tetrahydrofuranyl, and tetrahydropyranyl; in this class 4-piperidyl rings substituted with an aryl group at carbon 4 and on acyl or sulfonyl substituent at N1 are preferred.
In the case of optional substitution, one or more substituents may be present and a wide variety of substituents are possible. Preferred optional substituents for all compounds of the present invention are set out in the following list:
(a)xe2x80x94OH;
(b)xe2x80x94SH;
(c)xe2x80x94halogen, such as fluorine, chlorine or bromine,
(d)xe2x80x94CO2H;
(e)xe2x80x94CN;
(f)xe2x80x94NO2;
(g)xe2x80x94NR1R2 wherein each of R1 and R2 is selected from H and lower alkyl groups having 1 to 6 carbon atoms; or R1 and R2 together form a ring including 4 to 6 carbon atoms;
(h)xe2x80x94SO2NR1R2 where R1 and R2 are as defined above;
(i)xe2x80x94CONH2, xe2x80x94NHCO2H, or xe2x80x94NHCOCOOH;
(j) an alkyl (or alkenyl or alkynyl group) group having 1 to 12 (2 to 12) carbon atoms, preferably 1 to 7 (2 to 7) carbon atoms optionally substituted by any one or more of the groups (a)-(i) above and/or optionally interrupted by a group selected from xe2x80x94Oxe2x80x94, xe2x80x94Sxe2x80x94, xe2x80x94NR3xe2x80x94, 
xe2x80x94CO2xe2x80x94, xe2x80x94OCOxe2x80x94, xe2x80x94CONR3xe2x80x94, xe2x80x94NR3CONR3xe2x80x94, xe2x80x94SO2xe2x80x94, xe2x80x94NR3SO2xe2x80x94, and xe2x80x94SO2NR3xe2x80x94; where each R3 independently is H or lower alkyl of 1 to 6 carbon atoms;
(k) an aryl or heteroaryl group having 2 to 10 carbon atoms optionally substituted with any one or more of groups (a) to (j) above;
(l) an aralkyl or heteroaralkyl group having 3 to 16 carbon atoms optionally substituted with any one or more of groups (a)-(j) above and/or in which the alkyl part of the group is optionally interrupted by a group selected from xe2x80x94Oxe2x80x94, xe2x80x94Sxe2x80x94, xe2x80x94NR3xe2x80x94, 
xe2x80x94CO2xe2x80x94, xe2x80x94OCOxe2x80x94, xe2x80x94CONR3xe2x80x94, xe2x80x94NR3CONR3xe2x80x94, xe2x80x94SO2xe2x80x94, xe2x80x94NR3SO2xe2x80x94, and xe2x80x94SO2NR3xe2x80x94; where R3 is as defined above;
(m) 
where R4 is an alkyl, alkenyl, alkynyl, aryl, heteroaryl, aralkyl, or heteroaralkyl group as such groups are defined above at (j), (k) and (l);
(n) 
where R4 is as defined above;
(o) xe2x80x94OR4 where R4 is as defined above;
(p) 
where R4 is as defined above;
(q) xe2x80x94SO2R4 where R4 is as defined above;
(r) xe2x80x94NHR4 or xe2x80x94N(R4)2 where R4 is as defined above;
(s) xe2x80x94NHSO2R4 or xe2x80x94SO2NHR4, where R4 is as defined above;
(t) xe2x80x94SR4 
and each of optional substituents (j) to (t) above may optionally itself be substituted by one or more groups selected from (j) to (t).
A preferred class of compounds of formula A is represented by formula E: 
in which Ar is an optionally substituted aryl or heteroaryl group. Optional substituents may be selected from the list of preferred substituents set out above. Within this class of preferred compounds two especially preferred groups are set out below (formulas F and G) 
R5, R6, R7 and R8 are, independently H or are selected from the optional substituents listed above and R7 and R8 taken together may form a 4 to 7, preferably 5 or 6 membered ring; and X is O, S, NH, or NR4 where R4 is as defined above.
In compounds of formula F, (which are optionally substituted phenyl diketoacids) ortho, meta and para substitution are possible.
In general, it is preferred that there is a single substituent, preferably at the position which is ortho- or meta- to the diketoacid group. Substitution at the meta-position is especially preferred. Where two substituents are present, then preferably the phenyldiketoacid is 2,5-substituted; 3,5-substitution is also possible, as is 2,4-substitution provided, in the latter case, that the substituent at the 4-position is relatively small (e.g. methyl). Disubstitution at the 2,3- and 2,6-positions is, in general, not preferred.
Preferred substituents, especially at the ortho and meta positions, are ether groups of formula (o) above (i.e. xe2x80x94OR4), hydroxyl, and xe2x80x94NHSO2R4. It is generally preferred that no more than one substituent be xe2x80x94OR4 and/or xe2x80x94NHSO2R4.
Preferred examples of xe2x80x94OR4 groups which may be found at the ortho and meta positions and particularly at the meta position include:
xe2x80x94OCH2Ar or, less preferably xe2x80x94O(CH2)2Ar where Ar is an optionally substituted aryl or heteroaryl group and is particularly preferably an optionally substituted phenyl group. Examples of preferred substituents on the aryl group, and especially on the phenyl ring include halogens, especially fluorine and chlorine, and electron-withdrawing groups such as xe2x80x94CN, xe2x80x94CO2H, and xe2x80x94CF3 as well as ether and aryl groups;
xe2x80x94Oxe2x80x94(CH2)3xe2x80x94CN; and
xe2x80x94Oxe2x80x94(CH2)3xe2x80x94Cxe2x89xa1CH.
Preferred sulfonamide groups which may be found at the ortho- and meta- positions, particularly at the meta-position are those of formula:
xe2x80x94NHxe2x80x94SO2xe2x80x94Ar, where Ar is an optionally substituted aryl or heteroaryl group, preferably an optionally substituted phenyl group. Preferred optional substituents for the aryl, preferably phenyl group, include: xe2x80x94CN; halogens, especially chlorine and fluorine, xe2x80x94CF3, lower (C1-6) alkyl (especially methyl), hydroxy-, ether, and xe2x80x94NO2 groups.
For both the xe2x80x94OCH2Ar and xe2x80x94NHSO2Ar substituted compounds, another preferred example of Ar is naphthyl.
Other preferred substituents at the ortho and meta positions are lower (eg C1-6) alkyl groups, especially C1-4 alkyl, such as methyl and ethyl, but in particular methyl, aralkyl groups, especially phenylmethyl groups, optionally substituted in the phenyl ring, especially by a halogen, and nitrogen-containing substituents such as primary, secondary or tertiary amine groups, optionally in protonated form, amide, urethane, or urea groups in each of which examples there is a nitrogen atom bonded to the phenyl ring.
One particularly preferred sub class of compounds of formula F is those in which each of R5 and R6 is selected from H, HOxe2x80x94, R4Oxe2x80x94, and xe2x80x94NHSO2R4 provided that no more than one of R5 and R6 is R4Oxe2x80x94 or xe2x80x94NHSO2R4.
In compounds of formula G the diketoacid group may be at the 2- or 3- position of the ring. In many cases substitution at the 2-position is preferred.
Preferred examples of compounds of formula G are those in which the five membered aromatic ring, 
is a pyrrole or thiophene ring. In the case of the pyrrole-substituted diketoacids, the groups R7 and R8 may both be hydrogen and in many cases that is preferred. if R7 and R8 correspond to substituent groups, then these may be at any of the positions not already occupied by the diketoacid group. Examples of possible substituents include alkyl (especially methyl), halogen, and aralkyl (especially benzyl) groups.
One embodiment of pyrrole substituted diketoacid is that in which the diketoacid group is at the 2-position of the ring and where the only other substituent in the ring is on the nitrogen atom. In this case, preferred examples of the substituent R4 present on the nitrogen atom, include alkyl, aryl or aralkyl groups, particularly aralkyl (such as benzyl) groups. Where an aryl or aralkyl group is present these are preferably substituted with halogen atoms, such as fluorine or chlorine, or by cyano-groups.
In the case of the thiophene-substituted diketoacids a wide range of substituents R7 and R8 may be employed in various positions as will be evident from the tables infra. Preferred thiophenes have an aralkyl (such as optionally substituted benzyl) or aryl (such as optionally substituted phenyl) substituent, e.g. at the 5-position of the thiophene ring.
Compounds containing furanyl rings may also be useful, especially for inhibiting HIV reverse transcriptase.
Preferred substituents are optionally substituted aryl groups (especially optionally substituted phenyl). Substitution is preferably at the 5-position of the ring.
The formulae of numerous preferred specific compounds of the present invention are presented later below.
The compounds of the present invention having formula A may be prepared by a process which comprises reaction of a compound of formula B with a dialkyloxalate of formula C followed by hydrolysis of the resulting diketo-ester of formula D: 
where Rxe2x80x2 is an alkyl group, typically having 1-6 carbon atoms. In the case where the target molecule is a pharmaceutically acceptable ester of the compound of formula A then Rxe2x80x2 in formula C may be selected accordingly, and the step of hydrolysing the compound of formula D omitted, since in vivo hydrolysis can render the compounds active.
Preferred enzymes for inhibition by the compounds of the invention are those involved in phosphate transfer, in particular polymerases such as DNA polymerases, and RNA polymerases both of which may be either RNA dependent or DNA dependant. Compounds of the invention may particularly preferably be employed in the inhibition of viral enzymes. Examples of viral enzymes include RNA-dependent RNA polymerase and reverse transcriptases.
The compounds of the invention may be used as inhibitors of plant or animal (including human) viruses.
The viruses may be RNA viruses, which may, for example, be positive single stranded viruses of which polio virus, hepatitis C virus and encephalomyocarditis are examples, negative single stranded viruses such as orthomyxoviruses, and paramyxoviruses, and retroviruses of which HIV is a prominent example. Alternatively, the viruses may be DNA viruses, especially double stranded DNA viruses such as hepatitis B virus. In particular, compounds of the present invention may inhibit one or more of the following enzymes: hepatitis C virus RNA dependent RNA polymerase (HCV RdRp), hepatitis B virus polymerase (HBV pol) and reverse transcriptase of human immunodeficiency virus (HIV RT).
Especially preferred compounds of the invention will be suitable for use as HCV RdRp inhibitors.
Other classes of enzyme involved in phosphate transfer which may be susceptible to inhibition by compounds of the present invention include phosphatases, Rnases, integrases and ribozymes.
According to a further aspect of the invention there is provided the non-therapeutic use of compound of formula A or suitable salt or ester as an enzyme inhibitor, especially as an inhibitor of polymerases, especially viral polymerases. For instance, compounds of the invention may be of utility in agriculture and horticulture for treating plants infected with or susceptible to plant virus.
According to a further aspect of the invention there is provided the use of a compound of formula A or of a pharmaceutically acceptable salt or ester thereof in the manufacture of a medicament for treatment of a viral illness in a human or animal. For instance, the medicament may be used to treat viral illness by inhibiting one or more viral polymerase. Preferably the medicament is for treatment of hepatitis, such as hepatitis B or C, particularly hepatitis C, and human immunodeficiency virus.
A still further aspect of the invention provides a pharmaceutical composition comprising a compound of formula A, or a pharmaceutically acceptable salt or ester thereof and a pharmaceutically acceptable excipient, diluent or carrier. The composition may be in any suitable form, depending on the intended method of administration. It may for example be in the form of a tablet, capsule or liquid for oral administration, or of a solution or suspension for administration parenterally.
The pharmaceutical compositions optionally also include one or more other agents for the treatment of viral infections such as an antiviral agent, or an immunomodulatory agent such as xcex1-, xcex2-, or xcex3-interferon.
A still further aspect of the invention provides a method of inhibiting an enzyme, especially a viral polymerase and/or of treating or preventing a viral illness, the method involving administering to a human or animal (preferably mammalian) subject suffering from the condition a therapeutically or prophylactically effective amount of the pharmaceutical composition described above or of a compound of formula A or salt or ester thereof. xe2x80x9cEffective amountxe2x80x9d means an amount sufficient to cause a benefit to the subject or at least to cause a change in the subject""s condition.
The dosage rate at which the compound, salt or ester is administered will depend on the nature of the subject, the nature and severity of the condition, the administration method,used, etc. Appropriate values are selectable by routine testing. The compound, salt or ester may be administered alone or in combination with other treatments, either simultaneously or sequentially. For instance, it may be administered in combination with effective amounts of antiviral agents, immunomodulators, anti-infectives, or vaccines known to those of ordinary skill in the art. It may be administered by any suitable route, including orally, intravenously, cutaneously, subcutaneously, etc. It may be administered directly to a suitable site or in a manner in which it targets a particular site, such as a certain type of cell. Suitable targeting methods are already known.
A further aspect of the invention provides a method of preparation of a pharmaceutical composition, involving admixing one or more compound of formula A or salt or ester thereof with one or more pharmaceutically acceptable adjuvants, diluents or carriers and/or with one or more other therapeutically or prophylactically active agents.
Modes for Carrying Out the Invention
Embodiments of the invention are described below by the way of example only.